Fig. 4

(a–c) Chd protein distribution detected by confocal microscopy at different stages. Embryos injected with Tg(-2.067gsc:tBr-gfp) or Tg(-2.272bmp2b:tBr-gfp) DNA were harvested along with uninjected embryos at indicated stages for immunostaining using an anti-Chordin antibody and DAPI staining. Representative z-stack images were shown in animal-pole view with dorsal to the right. (d–g) The average ratios (e–g) of Chd to DAPI signals at the ventralmost or dorsalmost areas on optical equatorial sections at the blastodermal margin as illustrated in d. The strategy for estimating relative intensity was the same as described in the legend of Supplementary Fig. 2. For each group, the average relative immunostaining intensity was calculated from 3 to 5 embryos and three optical sections at and nearby the blastodermal margin per embryo. The error bars represented s.d. The Chd/DAPI ratio in the ventralmost region at each stage was set to 1.0. Statistical significance (Student’s t-test): NS, nonsignificant (P>0.05); *P<0.05; **P<0.01. (h–k) chd transcripts distribution detected by in situ hybridization at indicated stages. Representative embryos were animal-pole views with dorsal to the right. The ratio of embryos with representative pattern was indicated for those shown in h–j (l) The ratio of embryos showing an expansion of chd expression at the shield stage (as shown in k) n, number of observed embryos.